This invention was made with the assistance of Grant No. AM-30759 awarded by the National Institute of Health. The Government has certain rights in this invention.
It has been established that phosphate (Pi) is reabsorbed from luminal fluid of renal proximal tubules via secondary active, Na.sup.+ -gradient [Na.sup.+ extravesicular &gt;Na.sup.+ intravesicular; na.sup.+.sub.o &gt;] dependent uptake across the microvillar brush border membrane (BBM). Numerous properties of this renal secondary active transport of Pi, namely functional changes in response to hormones, drugs and nutritional stimuli have been recently described, but the molecular structure of the Na-Pi cotransporter within renal BBM remains unknown. A similar Na.sup.+ -gradient dependent transport has been stablished in BBM of the epithelium of the small intestine.
The usefulness of compounds currently known to inhibit the Na.sup.+ -gradient-dependent Pi transport across BBM is rather limited. Arsenate (AsO.sub.2.sup.+ 2) competitively inhibits the BBM transport system for Pi, albeit with relatively low affinity. However, arsenate also interacts with and inhibits another component of BBM, alkaline phosphatase, and interferes with transepithelial Pi reabsorption by uncoupling oxidative mitochondrial respiration of proximal tubules. The inhibitory effect of arsenate on intermediary metabolism limits its use as specific blocker of BBM transport of Pi in studies on the intact cells, and prevents its use in vivo. Nicotinamide adenine dinucleotide (NAD) binds on BBM and inhibits competitively the Na.sup.+ - gradient dependent Pi uptakes in vitro. However, NAD also inhibits renal BBM-bound alkaline phosphatase and, after prolonged incubation with BBM in vitro, NAD is catabolized by NAD glycohydrolase and is hydrolyzed to adenosine, Pi and other components.